Keratan sulfate proteoglycan is one of the major component of the corneal stroma, which composes about 90% of the total corneal thickness, and is important to maintain corneal transparency. Macular corneal dystrophy (MCD) is a hereditary eye disease, of which the patients show spotted opacity in their corneal stroma. The patients eventually need to be subjected to keratoplasty because of blindness by cornea clouding. Based on the studies of a carbohydrate sulfotransferase gene, CHST6, which is responsible for MCD, and the functional analysis of the gene product named human corneal GlcNAc 6-O sulfotransferase (hCGn6ST), we concluded that hCGn6ST is involved in the sulfation of keratan sulfate carbohydrate in the cornea. From these findings, we hypothesize that MCD phenotype is caused by the accumulation of unsulfated keratan sulfate proteoglycan, which is produced by the corneal cells lacking hCGn6ST activity, in the corneal tissue. We also hypothesize the presence of a corneal tissue-specific gene regulatory element(s) in CHST6 and mutations on the element result in the disruption of CHST6 expression in the corneal tissue. To address these issues, we propose the following specific aims; 1) To establish biosynthetic mechanism of corneal keratan sulfate 2) To identify the corneal tissue-specific gene regulatory element in CHST6 3) To produce the gene knockout mouse that is an animal model of MCD in human These results will provide us with information upon the involvement of keratan sulfate carbohydrate in the maintenance of the corneal transparency. Data obtained from this project are expected to form the basis for further clinical applications for MCD and the other corneal diseases through tissue engineering, medication and gene therapy.